Effect of cations on structure-linked sedimentability of lysosomal hydrolases.
1. A partially purified lysosomal preparation was obtained from mouse liver sucrose homogenates by differential and discontinuous gradient centrifugation. 2. Triton X-100 or repeated freezing and thawing of the lysosomal suspension (subfraction B) allowed comparison of free and activated values for acid phosphohydrolase, beta-glucuronidase and N-acetylglucosaminidase in the presence and absence of ascorbate. 3. The distribution of hydrolase activities between supernatant and pellet after high-speed centrifugation was measured and the percentages of total enzyme found in the supernatant were: acid phosphohydrolase, 40.7; beta-glucuronidase, 51; N-acetylglucosaminidase, 39.4. 4. Differential rates of elution of the three hydrolases from the membrane fraction occurred with increasing Na(+) and K(+) concentrations, whereas complex biphasic elution curves were obtained as a function of bivalent cation concentration with Ca(2+) and Mg(2+). 5. Sucrose-density-gradient centrifugation of frozen-and-thawed subfraction B demonstrated highly significant changes in the protein gradient profile in the presence of a low concentration of bivalent cation, indicating membrane aggregation and enzyme-membrane association. 6. The data provide further evidence for the nature of lysosomal enzyme binding and indicate the presence of different enzyme-membrane bonds conferring structure-linked latency upon individual lysosomal enzymes.
D000135:Acid Phosphatase / Q000032:analysis / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002499:Centrifugation, Density Gradient; D005966:Glucuronidase / Q000032:analysis / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000032:analysis / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology*; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D051379:Mice; D011188:Potassium / Q000494:pharmacology; D012964:Sodium; D013501:Surface-Active Agents
D013501:Surface-Active Agents; D012964:Sodium; D000135:Acid Phosphatase; D006026:Glycoside Hydrolases; D005966:Glucuronidase; D008274:Magnesium; D011188:Potassium; D002118:Calcium